Recently a gluconeogenic enzyme was discovered-fructose 1,6-bisphosphatase (FBPase)-that localizes in the nucleus of a proliferating cell, but its physiological role in this compartment remains unclear. Here, we demonstrate the link between nuclear localization of FBPase and the cell cycle progression. Results of our studies indicate that in human and mouse squamous cell lung cancer, as well as in the HL-1 cardiomyocytes, FBPase nuclear localization correlates with nuclear localization of S and G2 phase cyclins. Additionally, activity and expression of the enzyme depends on cell cycle stages. Identification of FBPase interacting partners with mass spectrometry reveals a set of nuclear proteins involved in cell cycle regulation, mRNA processing and in stabilization of genomic DNA structure. To our knowledge, this is the first experimental evidence that muscle FBPase is involved in cell cycle events.